Interpretive Summary: Transgenic technologies are powerful techniques for creating useful, genetically modified plants, also known as GMOs. Although there are many examples of widely used, and successful, GMO varieties, there are still problems and limitations of the technology that must be solved. For instance, selectable markers, such as antibiotic or herbicide resistance, are not viewed favorably by consumers and they also can lead to problems with the performance of the GMO crop. In this study, we found that the use of a transposon system to be effective for producing GMO barley that expresses the engineered trait very well. However, we also discovered that this good characteristic also causes a problem because the gene, a selectable marker gene for herbicide resistance, reduces the health and yield of the plants when it is expressed well. These results clearly demonstrate an undesirable effect of this herbicide resistance gene in barley, and provide guidance for producing superior GMO barley plants. Future work will now concentrate on using systems for producing GMO barley that do not contain selectable marker genes.

Technical Abstract:
Selectable markers used in plant transformation, such as phosphinothricin acetyltransferase (PAT) derived from the bar gene, have been chosen for selection efficacy as well as for the absence of pleiotropic effects. Recent research has suggested that expression of bar in Arabidopsis affects the transcriptome, and in at least one barley cultivar high bar expression has been associated with severe fitness reductions. However, the observed sensitivity in barley was thought to be an isolated case associated with a specific genetic stock. In this study, fitness reductions were observed in BC8 near-isogenic lines (NILs), in the background of the cultivar Garnet, when Ds-bar was brought to homozygosity. Fitness of NILs was correlated with the level of expression in donor parents. Previous studies had shown heritability of bar expression in F1 barley hybrids, so these results suggested that Garnet barley is intolerant of high PAT activity. PAT activities were determined for eight NILs selected that showed a range of phenotypes (apparently normal to abnormal and non-viable) in response to homozygosity for Ds-bar. Fitness was clearly and negatively correlated to PAT activities. These results suggest genetic variability for sensitivity to bar expression in barley, but cannot rule out the possibility that negative but non-obvious effects occur in apparently tolerant cultivars. Similar problems may be associated, but not yet detected, with other selectable markers. Thus, it is recommended that transformation of barley be conducted using strategies that enable the elimination of selectable markers.